It is planned to continue our studies on the factors controlling thyroid hormones transfer from blood to cells, their intracellular metabolism and finally their mode of action. A simple tissue culture model was chosen to achieve these goals. Hormone transfer from medium to cells and their subcellular distribution, with particular emphasis to nuclear binding, will be studied utilizing radioiodinated thyroid hormones. Metabolism of thyroid hormones possibly related to hormone action will be studied in human fibroblasts and rat pituitary cell lines, and correlated to the degradative metabolism in liver cell lines. The role of desialylated human thyroxine binding globulin on the hormonal uptake by liver will be also investigated. Indications for cellular action of thyroid hormone will be derived from its suppressive effect on the thyrotropin releasing hormone (TRH) mediated proclactin release in pituitary cells. The relative potency of thyroxine (T4) and triiodothyronine (T3) and the necessity of monodeiodination of T4 to T3 for hormone action will be also examined. Conversion of T4 to T3 will be blocked with prolythiouracil. Studies will be undertaken to correlate the interaction of T3 and TRH to their uptake, their respective binding to nuclei and plasma membranes, and effects on cyclic-AMP. A possible relationship between thyroid hormone binding to nuclear receptors and RNA synthesis will be evaluated. Analyses of messenger RNA will be made, taking advantage of the newly developed method for labeling RNA to high specific activity with radioiodide. Finally, these studies will be applied to a cell line derived from patients with resistance to the action of thyroid hormone in an attempt to ellucidate this inherited defect.